Human Cell Biology ELISA Kits 2
Human COL1 alpha2 (Collagen Type I Alpha 2) ELISA Kit (HUES01914)
- SKU:
- HUES01914
- Product Type:
- ELISA Kit
- Size:
- 96 Assays
- Uniprot:
- P08123
- Sensitivity:
- 0.09ng/mL
- Range:
- 0.16-10ng/mL
- ELISA Type:
- Sandwich
- Synonyms:
- COL1A2, OI4
- Reactivity:
- Human
- Sample Type:
- Serum, plasma and other biological fluids
- Research Area:
- Cell Biology
Description
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Human |
| Detection Method: | Colormetric |
| Detection Range: | 0.16-10 ng/mL |
| Sensitivity: | 0.10 ng/mL |
| Sample Volume Required Per Well: | 100µL |
| Sample Type: | Serum, plasma and other biological fluids |
| Specificity: | This kit recognizes Human COL1 alpha2 in samples. No significant cross-reactivity or interference between Human COL1 alpha2 and analogues was observed. |
This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Human COL1 alpha2. Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human COL1 alpha2 and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human COL1 alpha2, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Human COL1 alpha2. The concentration of Human COL1 alpha2 in samples can be calculated by comparing the OD of the samples to the standard curve.
| UniProt Protein Function: | COL1A2: Type I collagen is a member of group I collagen (fibrillar forming collagen). Defects in COL1A2 are the cause of Ehlers-Danlos syndrome type 7B (EDS7B). EDS is a connective tissue disorder characterized by hyperextensible skin, atrophic cutaneous scars due to tissue fragility and joint hyperlaxity. EDS7B is marked by bilateral congenital hip dislocation, hyperlaxity of the joints, and recurrent partial dislocations. Defects in COL1A2 are a cause of osteogenesis imperfecta type 1 (OI1). A dominantly inherited connective tissue disorder characterized by bone fragility and blue sclerae. Osteogenesis imperfecta type 1 is non-deforming with normal height or mild short stature, and no dentinogenesis imperfecta. Defects in COL1A2 are a cause of osteogenesis imperfecta type 2 (OI2); also known as osteogenesis imperfecta congenita (OIC) or lethal perinatal. A connective tissue disorder characterized by bone fragility, with many perinatal fractures, severe bowing of long bones, undermineralization, and death in the perinatal period due to respiratory insufficiency. Defects in COL1A2 are the cause of Ehlers-Danlos syndrome autosomal recessive cardiac valvular form (EDSCV). A connective tissue disorder characterized by hyperextensible skin, atrophic cutaneous scars due to tissue fragility and joint hyperlaxity. In addition to joint laxity, skin hyperextensibility and friability, and abnormal scar formation, patients have mitral valve prolapse and insufficiency, mitral regurgitation, and aortic insufficiency. Defects in COL1A2 are a cause of osteogenesis imperfecta type 3 (OI3). A connective tissue disorder characterized by progressively deforming bones, very short stature, a triangular face, severe scoliosis, grayish sclera, and dentinogenesis imperfecta. Defects in COL1A2 are a cause of osteogenesis imperfecta type 4 (OI4); also known as osteogenesis imperfecta with normal sclerae. A connective tissue disorder characterized by moderately short stature, mild to moderate scoliosis, grayish or white sclera and dentinogenesis imperfecta. A chromosomal aberration involving COL1A2 may be a cause of lipoblastomas, which are benign tumors resulting from transformation of adipocytes, usually diagnosed in children. Translocation t(7;8)(p22;q13) with PLAG1. Belongs to the fibrillar collagen family. |
| UniProt Protein Details: | Protein type:Secreted, signal peptide; Secreted Chromosomal Location of Human Ortholog: 7q22. 1 Cellular Component: collagen type I; endoplasmic reticulum lumen; extracellular matrix; extracellular region; extracellular space Molecular Function:extracellular matrix structural constituent; identical protein binding; metal ion binding; platelet-derived growth factor binding; protein binding; protein binding, bridging; SMAD binding Biological Process: blood coagulation; blood vessel development; collagen catabolic process; collagen fibril organization; extracellular matrix disassembly; extracellular matrix organization and biogenesis; leukocyte migration; odontogenesis; platelet activation; receptor-mediated endocytosis; regulation of blood pressure; regulation of immune response; Rho protein signal transduction; skeletal development; skin morphogenesis; transforming growth factor beta receptor signaling pathway Disease: Ehlers-danlos Syndrome, Autosomal Recessive, Cardiac Valvular Form; Ehlers-danlos Syndrome, Type Vii, Autosomal Dominant; Osteogenesis Imperfecta, Type Ii; Osteogenesis Imperfecta, Type Iii; Osteogenesis Imperfecta, Type Iv; Osteoporosis |
| NCBI Summary: | This gene encodes the pro-alpha2 chain of type I collagen whose triple helix comprises two alpha1 chains and one alpha2 chain. Type I is a fibril-forming collagen found in most connective tissues and is abundant in bone, cornea, dermis and tendon. Mutations in this gene are associated with osteogenesis imperfecta types I-IV, Ehlers-Danlos syndrome type VIIB, recessive Ehlers-Danlos syndrome Classical type, idiopathic osteoporosis, and atypical Marfan syndrome. Symptoms associated with mutations in this gene, however, tend to be less severe than mutations in the gene for the alpha1 chain of type I collagen (COL1A1) reflecting the different role of alpha2 chains in matrix integrity. Three transcripts, resulting from the use of alternate polyadenylation signals, have been identified for this gene. [provided by R. Dalgleish, Feb 2008] |
| UniProt Code: | P08123 |
| NCBI GenInfo Identifier: | 296439507 |
| NCBI Gene ID: | 1278 |
| NCBI Accession: | P08123. 7 |
| UniProt Secondary Accession: | P08123,P02464, Q13897, Q13997, Q13998, Q14038, Q14057 Q15177, Q15947, Q16480, Q16511, Q7Z5S6, |
| UniProt Related Accession: | P08123 |
| Molecular Weight: | 129,314 Da |
| NCBI Full Name: | Collagen alpha-2(I) chain |
| NCBI Synonym Full Names: | collagen type I alpha 2 |
| NCBI Official Symbol: | COL1A2 |
| NCBI Official Synonym Symbols: | OI4 |
| NCBI Protein Information: | collagen alpha-2(I) chain |
| UniProt Protein Name: | Collagen alpha-2(I) chain |
| UniProt Synonym Protein Names: | Alpha-2 type I collagen |
| UniProt Gene Name: | COL1A2 |
| UniProt Entry Name: | CO1A2_HUMAN |
As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e. g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
| Concentration (ng/mL) | O.D | Average | Corrected |
| 10 | 2.431 2.445 | 2.438 | 2.37 |
| 5 | 1.525 1.561 | 1.543 | 1.475 |
| 2.5 | 0.907 0.891 | 0.899 | 0.831 |
| 1.25 | 0.41 0.418 | 0.414 | 0.346 |
| 0.63 | 0.28 0.266 | 0.273 | 0.205 |
| 0.32 | 0.169 0.165 | 0.167 | 0.099 |
| 0.16 | 0.116 0.122 | 0.119 | 0.051 |
| 0 | 0.067 0.069 | 0.068 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human COL1 alpha2 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human COL1 alpha2 were tested on 3 different plates, 20 replicates in each plate.
| Intra-assay Precision | Inter-assay Precision | |||||
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean (ng/mL) | 0.52 | 1.59 | 4.86 | 0.56 | 1.45 | 5.11 |
| Standard deviation | 0.03 | 0.07 | 0.15 | 0.03 | 0.07 | 0.26 |
| C V (%) | 5.77 | 4.40 | 3.09 | 5.36 | 4.83 | 5.09 |
Recovery
The recovery of Human COL1 alpha2 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
| Sample Type | Range (%) | Average Recovery (%) |
| Serum (n=5) | 89-103 | 94 |
| EDTA plasma (n=5) | 87-100 | 92 |
| Cell culture media (n=5) | 87-102 | 93 |
Linearity
Samples were spiked with high concentrations of Human COL1 alpha2 and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
| Serum (n=5) | EDTA plasma (n=5) | Cell culture media (n=5) | ||
| 1:2 | Range (%) | 100-114 | 99-112 | 93-108 |
| Average (%) | 105 | 105 | 100 | |
| 1:4 | Range (%) | 86-100 | 83-96 | 87-101 |
| Average (%) | 92 | 88 | 93 | |
| 1:8 | Range (%) | 90-101 | 85-100 | 84-96 |
| Average (%) | 96 | 92 | 91 | |
| 1:16 | Range (%) | 89-104 | 83-95 | 84-97 |
| Average (%) | 96 | 88 | 91 |
An unopened kit can be stored at 4°C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.
| Item | Specifications | Storage |
| Micro ELISA Plate(Dismountable) | 8 wells ×12 strips | -20°C, 6 months |
| Reference Standard | 2 vials | |
| Concentrated Biotinylated Detection Ab (100×) | 1 vial, 120 µL | |
| Concentrated HRP Conjugate (100×) | 1 vial, 120 µL | -20°C(shading light), 6 months |
| Reference Standard & Sample Diluent | 1 vial, 20 mL | 4°C, 6 months |
| Biotinylated Detection Ab Diluent | 1 vial, 14 mL | |
| HRP Conjugate Diluent | 1 vial, 14 mL | |
| Concentrated Wash Buffer (25×) | 1 vial, 30 mL | |
| Substrate Reagent | 1 vial, 10 mL | 4°C(shading light) |
| Stop Solution | 1 vial, 10 mL | 4°C |
| Plate Sealer | 5 pieces | |
| Product Description | 1 copy | |
| Certificate of Analysis | 1 copy |
- Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
- Aliquot 100 µL of standard solutions into the standard wells.
- Add 100 µL of Sample / Standard dilution buffer into the control (zero) well.
- Add 100 µL of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- Cover the plate with the sealer provided in the kit and incubate for 90 min at 37 °C.
- Aspirate the liquid from each well, do not wash. Immediately add 100 µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix. Incubate for 1 hour at 37 °C.
- Aspirate or decant the solution from the plate and add 350 µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
- Add 100 µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37 °C.
- Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- Add 90 µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37 °C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
- Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.
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